G-protein coupled receptors (GPCRs) are one of the most biologically and clinically important class of proteins, representing greater than 1% of the vertebrate genomes and greater than 50% of all pharmaceutical targets. Intriguingly, the activating ligands for these receptors range from full proteins to photons, yet we are unable to detail the structure-function relationships for GPCRs because of the great difficulty in generating high resolution crystal structures of these seven-transmembrane receptors. Combinatorial mutagenesis strategies will be employed to directly address two of the biggest limitations in producing GPCRs suitable for crystallographic studies, low expression/stability and inadequate diffraction quality of receptor crystals. First, error-prone polymerase chain reaction methods will be used to generate a receptor library that can be transformed into bacteria and screened for expression level using a selection marker such as a fluorescent protein fusion or an essential enzyme. This smaller library can then by solubilized in detergent solution and screened against immobilized ligand, thereby testing for receptor stability out of the membrane as well as competency in ligand binding. Second, ribosome display will be used to select novel ankyrin-repeat proteins that bind to specific epitopes on GPCRs. Such moieties may be used as adjuvants for GPCR crystallization, since ankyrin-repeat proteins, which readily crystallize, may induce a co-crystal lattice formation that has a greatly improved diffraction quality. This work will have a wide-reaching impact, ranging from fundamental studies on receptor biology and signal transduction to clinical applications in structure-based drug design. [unreadable] [unreadable]